7-heterocyclic-substituted-acylamido cephalosporins



United States Patent M 3,218,318 7-HETEROCYCLlC-SUBSTITUTED-ACYLAMIDO CEPHALOSPORINS Edwin H. Flynn, Indianapolis, lind., assignor to Eli Lilly and Company, Indianapolis, Ind., a corporation of Indiana No Drawing. Filed Aug. 31, 1962, Ser. No. 220,855 8 Claims. (Cl. 260-243) This application is a continuation-in-part of my application Serial No. 115,612, filed June 8, 1961, now abandoned.

This invention relates to novel organic compounds and to methods for their preparation.

The novel compounds of the present invention are heteromonocyclic-substituted acyl derivatives of the nuclei of compounds of the class of cephalosporin C, having the heteromonocyclic-substituted acylamido group in the 7 position of the cephalosporin C compound instead of the '-amino-N-adipamyl group. The novel compounds are represented by the following formula:

in which R taken alone, is -OH, C C acyloxy, or tertiary-amino, R is -OH when R is -OH, R is OH when R is C -C acyloxy, R is -O when R is tertiary-amino, R and R when taken together, are O, n is Zero or 1, R is C -C alkylene, and R is a heteromonocyclic radical containing 0, S, and/ or N.

Thus, R can be acetoxy, propionoxy, butyroxy, capryloxy or the like; or N-pyridyl, N-pyrimidyl, trimethylaniino, triethylamino, tributylamino, or other tertiary-amino group such as those produced by reaction of cephalosporin C with nicotine, nicotinic acid, isonicotinic acid, nicotinamide, Z-aminopyridine, 2-amino-6-methy1pyridine, 2,4,6- trimethylpyridine, 2 hydroxymethylpyridine, sulfapyridine, 3-hydroxypyridine, pyridine-2,3-dicarboxylic acid, quinoline, sulfadiazine, sulfathiazole, picolinic acid, and the like.

R can be methylene, dimethylene, trimethylene, tetramethylene, pentamethylene, hexamethylene, and the methyl, dimethyl, ethyl, diethyl, and other alkyl substitution products thereof containing a maximum of six carbon atoms.

R, can be dioxanyl, Z-furyl, 3-furyl, imidazolyl, isoxazolyl, morpholinyl, oxazolyl, pyranyl, pyrazinyl, pyrazolyl, N-pyridyl, Z-pyridyl, 3-pyridyl, pyrimidyl, N-pyrryl, Z-pyrryl, 3-pyrryl, thiazolyl, 2-thienyl, 3-thienyl, triazinyl, triazolyl, and the like; the partially and completely hydrogenated derivatives thereof such as tetrahydrofuryl, imidazolinyl imidazolinyl, imidazolidyl, piperidyl, tetrahydropyrimidyl, pyrrolidyl, and the like; and the derivatives thereof having chlorine, fluorine, bromine, iodine, nitro, ,methoxy, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, or other C -C alkyl substituent at one or more positions on the ring, e.g., the picolyls, the methylfuryls, the methylthienyls, nitrofuryl, nitropyridyl, nitrothienyl, chlorofuryl, bromopyridyl, fiuoropyrryl, methoxypyrimidyl, and the like.

The novel compounds of the present invention are related to cephalosporin C insofar as they contain the 2,3- dihydro-6I-I-1,5-thiazine ring with a fused [i-lactam ring in the 5,6 position, which is characteristic of cephalosporin C. However, unlike cephalosporin C, which contains the 5'-amino-N-adipamyl group in the 7 position,

3,218,318 Patented Nov. 16, 1965 the compounds of the present invention are characterized by a heteromonocyclic-substituted acylamido group in the 7 position. Moreover, unlike cephalosporin C, which has a relatively low antibacterial action, the compounds of the present invention are highly effective antibacterial agents. They are further characterized by penicillinase-resistance, acid stability, and activity against a broad range of microorganisms, including both Gram-positive and Gram-negative pathogens.

As will be observed from the formulas given above, the invention includes a variety of related compounds having the bicyclic ring structure of cephalosporin C, but with variations in the substituent groups attached thereto. Among such compounds are those having the nuclei of the cephalosporin-type products known as cephalosporin C desacetylcephalosporin C, and cephalosporin C these nuclei being represented by the following formulas, respectively:

/C I 2 Am+ toowhere Arn represents a tertiary-amino radical, exemplified above. As will be seen from the above formulas, the nucleus of cephalosporin C includes a fused lactone ring, while the nucleus of cephalosporin C forms an inner salt or zwitterion.

In general, those compounds which possess the lactone type nucleus (i.e., the C nucleus) are less active antibacterially than those having the cephalosporin C nucleus. Also in general, those compounds which possess the cephalosporin C nucleus are more effective antibacterially than those containing the cephalosporin C nucleus. However, this differentiation is only of general application, and examples are to be found in which reverse activities exist.

As is the case with the penicillins to which the compounds of this invention are in some degree related, numerous salts, esters, amides, and the like derivativesthereof can be prepared by combination with nontoxic pharmaceutically acceptable cations, anions, alcohol residues, ammonia, and amines, and such derivatives are to be regarded as the full equivalents of the compounds disclosed and claimed herein, and accordingly are to be considered as within the scope of this invention.

For purpose of illustration, there can be mentioned several types of cationic salts which can be prepared from compounds containing the cephalosporin C nucleus, including, for example, water-soluble salts such as the sodium, potassium, lithium, ammonium, and substituted ammonium salts, as well as the less water-soluble salts such as the calcium, barium, procaine, quinine, and dibenzylethylenediamine salts. Those compounds which contain the cephalosporin C nucleus do not form cationic salts but instead form anionic salts, i.e., acid addition salts, with strong acids such as hydrochloric, hydrobromic, phosphoric, sulfuric, and like acids.

The following examples, together with the operating examples appearing hereinafter, will illustrate the types of compounds available in accordance with the present invention:

7- (2'-pyranylacetamido cephalosporanic acid 7- 2'-piperidylacetamido) cephalosporanic acid 7- 2'-pyridylacetamido cephalosporanic acid 7- 'oxazolylacetamido) cephalosporanic acid 7- (4( 5 -imidazolylacetamido) cephalosporanic acid 7- 2'-morpholinylacetamido cephalosporanic acid 7- 2-pyrazinylacetamid-o cephalosporanic acid 7- 5 '-imidazolinylacetamido cephalosp oranic acid 7- 3-bromo-2'-pyridylacetamido cephalosporanic acid 7 8 5 '-fluoro-2-pyridyl propionamidocephalosporanic acid 7-,8- 5 -pyrimidyl propionamidocephalosporanic acid 7- 5 nitro-2'-thienylacetamido) cephalosporanic acid 7- 2'-thiazolylacetamido) cephalosporanic acid 7-'y-(2-furyl)butyramidocephalosporanic acid 7-6-(2'-thienyl) valeramidocephalosporanic acid 7-e- 3 '-thienyl hexanoylamidocephalosporanic acid 7- OL-PlCOlYl-3 '-acetamido cephalosporanic acid 7-a-2'-thienylpropionamidocephalosporanic acid 7-a-2-thienylisobutyramidocephalosporanic acid 7-u-2'furyl-n-butyramidocephalosporanic acid 7-( 2-methyl-3-furylacetamido cephalosporanic acid 7- 3 '-methyl-2-thienylacetamido cephalosp oranic acid 7- 5 -methoxy-3-pyridylacetamido cephalosporanic acid 7- (2-triazinylacetamido cephalosporanic acid and the like, including the cephalosporin C and cephalosporin C analogues thereof.

Cephalosporin C can be prepared by cultivating a cephalosporin C-producing organism in a suitable nutrient medium, as described in British patent specification 810,196, published March 11, 1959.

Cephalosporin C is readily converted into cephalosporin C by heating with water under acid conditions, as described in Belgian Patent 593,777, published November 30, 1960. This removes the acetyl group from its point of attachment through oxygen to the methyl group in the 3 position of the thiazine ring, and lactonization then spontaneously occurs, yielding the fused cyclic lactone.

Cephalosporin C is also readily converted into compounds of the cephalosporin C type by refluxing in aqueous solution with an excess of pyridine, for example, as described in Belgian Patent 593,777. The reaction is applicable in general to the tertiary amines, of which numerous examples are given above, yielding corresponding derivatives of the cephalosporin C type wherein the tertiary amine is attached to the methyl group in the 3 position of the thiazine ring, and forms an inner salt with the carboxyl group in the 4 position.

Desacetylcephalosporin C is conveniently prepared by treating cephalosporin C with citrus acetylesterase for several hours in aqueous phosphate buffer at pH 6.5-7 according to the method of Jansen, Jang, and MacDonnell, Archiv. Biochem., 15 (1947), 415-31.

From the various cephalosporin C compounds thus available, the corresponding nucleus is readily obtained by cleaving the 5-amino-N'-adipamyl side chain between its amido nitrogen and its amido carbonyl group. Thus, 7-aminocephalosporanic acid can be obtained by digesting cephalosporin C for an extended period in the presence of a mineral acid and in the absence of light, according the conversion procedures of Belgian Patent 593,777 and of Jansen et al. to produce compounds having the respective nuclei.

For the acylation of the 7-amino group of the cephalosporin nucleus, as defined above, any of the conventional acylation procedures can be employed, utilizing any of the various types of known acylating agents having a composition which yields the desired side chain.

A convenient acylating agent is the appropriate heteromonocyclic-substituted acyl chloride or bromide. The acylation is carried out in water or an appropriate organic solvent, preferably under substantially neutral conditions, and preferably at reduced temperature, i.e., above the freezing point of the reaction mixture and up to about 20 C. In a typical procedure, 7-aminocephalosporanic acid or one of its derivatives as defined herein, together with a sufficient quantity of sodium bicarbonate or other appropriate alkali to promote solution, is dissolved in aqueous 50 volume-percent acetone, the concentration of the 7-aminocephalosporanic acid being about 1 to about 4 percent by weight. The solution is cooled to around 0 to 5 C., and a solution of the acylating agent is added in about 20 percent excess, with stirring and cooling. The pH of the mixture can be maintained, if it tends to vary, around the neutral level by bubbling carbon dioxide therein. After addition of the acylating agent has been completed, stirring of the reaction mixture is continued, and the mixture is allowed to warm to room temperature. The reaction product is then acidified to around pH 2 and extracted with an organic solvent such as ethyl acetate. The ethyl acetate extract is adjusted to around pH 5.5 with a base containing the desired cation of the final product, and is extracted with water. The water solution is separated and evaporated to dryness. The residue is taken up in the minimum quantity of water, and the desired product is precipitated by adding a large excess of acetone and, if necessary, ether. The crystalline product obtained thereby is filtered, washed with acetone, and dried.

Acylation can also be carried out with the corresponding heterornonocyclic-substituted alkanoic acids, employed in conjunction with an equimolar proportion of a carbodiimide such as N,N'-diisopropylcarbodiimide, N,N-dicyclohexylcarbodiimide, N,N-bis(p-dimethylaminophen yl)carbodiimide, or the like, and the acylation proceeds at ordinary temperatures in such cases. Alternatively, the heteromonocyclic-substituted alkanoic acids can be converted into the corresponding acid anhydride, or into the azide, or into an activated ester, and any of these derivatives can be used to effect the desired acylation. Other agents can readily be ascertained from the art.

Many of the acylating agents, together with methods for their preparation, are described in the literature, and a number of them are commercially available. All of them are readily prepared by methods well known in the art.

The invention will be more readily understood from the following operating examples, which are submitted as illustrations only, and not by way of limitation.

EXAMPLE 1 7- (2'-fur0ylamid0) cephalosporanic acid 7-Aminocephalosporanic acid (1.0 g.) and sodium bicarbonate (ca. 1 g.) were dissolved in a mixture of 50 ml. of water and 40 ml. of acetone. The solution was stirred .in an ice bath and to it was added 450 mg. of 2-furoyl chloride (commercially available) dissolved in 10 ml. of acetone over a period of about 30 minutes, after which the mixture was stirred around 2 /2 additional hours in the cold. The reaction product mixture was then stripped of acetone under vacuum, and ml. of ethyl acetate were added, followed by 1 N hydrochloric acid to pH 2. The aqueous phase was separated, washed with 50 ml. of

ethyl acetate, and discarded. The ethyl acetate layers were combined and washed with 50 ml. of water. The washed ethyl acetate extract was stirred with 100 ml. of water and adjusted to pH 5.5 with aqueous 0.5 N potassium hydroxide solution. The resulting aqueous extract was separated and evaporated to dryness under vacuum. The residue was triturated with aqueous acetone, and the solids were filtered oil and dried under vacuum. The yield was 870 mg. of 7-(2-furoylamido)cephalosporanic acid in the form of the potassium salt, having a maximum in its ultraviolet spectrum at 257 m (e=19,000).

The following additional compositions were prepared, employing methods and conditions generally approximating the foregoing:

EXAMPLE 2 7-[3-(2'-furyl)propionamidocephalosporanic acid potassium salt, having a maximum in its ultraviolet absorption spectrum at 260 m (e=7350). Yield, 980 mg. of product from 590 mg. of fi-(Z-furyDpropionyl chloride, prepared from B-(2-furyl)propionic acid (Dunlop, The Furans, New York, Reinhold, 1952, p. 588) by the following procedure:

,B-(2-furyl)propionic acid (1.4 g.) was dissolved with sodium bicarbonate (0.84 g.) in 20 ml. of water, and the solution was evaporated to dryness under vacuum. The residue was powdered in a mortar and dried in an oven overnight. The dry powder was slurried in benzene and cooled in an ice bath, and to the slurry were added two drops of pyridine and 3.8 g. of oxalyl chloride. The reaction began immediately. After one hour, the ice bath was removed and stirring was continued for two hours while the reaction mixture warmed to room temperature. The mixture was evaporated to dryness under vacuum. The residue was triturated with benzene, stripped again, and triturated again with benzene. The solution was filtered to remove inorganic solids. The filtrate was stripped under vacuum to a light-colored liquid, comprising largely fl-(2-furyl)propionyl chloride. This substance is unstable, and was therefore used promptly for the desired acylation.

EXAMPLE 3 Example 2.

EXAMPLE 4 7-(2'-thienylacetamido)cephalosporanic acid potassium salt, having a maximum in its ultraviolet absorption spectrum at 236 mp. (e=1l,500). Yield, 1.04 g. of product, from 560 mg. of 2-thienylacetyl chloride, obtainable by treatment of 2-thienylacetic acid (Ernst, Berichte, 19 (1886), 3281) with thionyl chloride in a conventional manner.

EXAMPLE 5 7-(3'-thienylacetamido)cephalosporanic acid potassium salt, lustrous needles upon crystallization from water, with maxima in the ultraviolet absorption spectrum at 236 m, (e=10,l70) and 260 m (e=7,240). Prepared from 470 mg. of 3-thienylacetyl chloride, obtained from 3-thienylacetic acid (Campaigne and Le Suer, I. Am. Chem. Soc., 70 (1948), 1555 by treatment with thionyl chloride in a conventional manner.

EXAMPLE 6 7-('y-2-thienyl-n-butyramido)cephalosporanic acid potassium salt, from 695 mg. of -Z-thienyI-n-butyryl chloride. Recrystallization of the crude product from methyl and isopropyl alcohols yielded 650 mg. of purified material having a maximum in its ultraviolet absorption spectrum at 235 m (e=11,750) and a shoulder at 260 m (e=7,700). The acyl chloride was prepared by treatment of the corresponding acid (Aldrich Chemical Company, Milwaukee, Wisconsin), with thionyl chloride in a conventional manner.

EXAMPLE 7 7-(3-methyl-2'-thienyl)acetamidocephalosporanic acid potassium salt, 610 mg., from 630 mg. of 3-methyl-2-thienylacetyl chloride. The product had a maximum in its ultraviolet absorption spectrum at 243 m (e=ll,420). The acyl chloride was prepared by the method of Hartough et al., J. Am. Chem. Soc., 69 (1947), 3093.

EXAMPLE 8 7- 5 -t-butyl-2'-thienyl acetamidocephalosporanic acid potassium salt, from 5-t-butyl-2-thienylacetyl chloride. The product, after crystallization from methyl and isopropyl alcohols, weighed 850 mg. and had a maximum in its ultraviolet absorption spectrum at 243 m (e=l3,700). The acyl chloride (800 mg.) was obtained from 5-t-butyl-2-thienylacetic acid (Prescott et al. J. Am. Chem. Soc., 72 (1950), 2110 at 2111) by treatment with thionyl chloride under conventional conditions.

EXAMPLE 9 7 (1'-methyl-4-nitropyrrole-2-carboxamido)cephalosporanic acid potassium salt, 600 mg., from 1-methyl-4- nitropyrryl-Z-carboxylic acid chloride, 700 mg. The product had a maximum in its ultraviolet absorption spectrum at 246 m (e=22,200), a pK of 4.78, a molecular weight of 458 (theo., 462), and an infrared absorption spectrum consistent with the expected structure. The pyrrolecarboxylic acid chloride was prepared by the method of Waller et al., U.S. Pat. 2,785,182 (March 12, 1957).

7- (1 -pyraz0leacezamid0 cephalosporanic acid To a solution of l-pyrazoleacetic acid (1.26 g., prepared as described in J. Org. Chem., 19 (1954), 1432) dissolved in 50 ml. of tetrahydrofuran was added dropwise a solution of 2.1 g. of dicyclohexylcarbodiimide in 50 ml. of tetrahydrofuran, followed by a solution of 2.72 g. of 7-aminocephalosporanic acid in 10 ml. of water, dissolution of the latter having been effected by adding a minimum amount of triethylamine. The resulting mixture was stirred overnight, then filtered, and the filtrate was evaporated to dryness under vacuum.

To the residue were added ml. of water and 100 ml. of ethyl acetate, and the mixture was adjusted to pH 2 with 10 percent hydrochloric acid. A small quantity of solid was filtered off and discarded. The layers were separated. The aqueous layer was washed with ethyl acetate and discarded. The ethyl acetate layers were combined and adjusted to pH 5.5 with aqueous 1 N potassium hydroxide. The layers were separated, and the aqueous layer was evaporated to dryness under vacuum. The solids obtained thereby were dissolved in methanol, diluted with isopropyl alcohol, and evaporated to somewhat less than half the original volume of the methanol solution. The resulting solids were filtered off and dried. The yield was 1.03 g. of 7-(1'-pyrazoleacetamido)cephalosporanic acid in the form of the potassium salt, having a maximum in its ultraviolet absorption spectrum at 260 III u. (6:7700

EXAMPLE 12 thiazole-S-acetic acid (Burger et al., J. Org. Chem., 12

(1947), 342 at 350). The product had a maximum in its ultraviolet absorption spectrum at 254 m (e=10,750).

EXAMPLE 14 7-(N'-methyl-2-pyrryl)acetamidocephalosporanic acid A solution of 1.5 g. of dicyclohexylcarbodiimide in v25 ml. of tetrahydrofuran was added to 1.0 g. of l-methyl- 2-pyrroleacetic acid (Nenitzescu et al., Berichte, 64 (1931), 1927) in 25 ml. of tetrahydrofuran. To the resulting mixture was added a solution of 7-aminocephalosporanic acid, prepared by commingling 2.0 g. of 7-aminocephalosporanic acid with ml. of water and adding triethylamine to pH 7, and the total mixture was stirred overnight.

The reaction product mixture was filtered and the tetrahydrofuran was stripped out under vacuum. The residual aqueous solution was stirred with 100 ml. of water and 100 ml. of ethyl acetate and adjusted to pH 2.0 with 1 N hydrochloric acid. The ethyl acetate phase was separated and adjusted to pH 6.5 with 50 ml. of water and a sufiicient quantity of aqueous 1 N potassium hydroxide solution. The aqueous phase was separated and evaporated to dryness under vacuum. The residue was triturated with a mixture of methyl and isopropyl alcohol-s. The resulting solid was filtered oh and dried. Yield, 650 mg. of 7-(N'-methyl-2-pyrryl)acetamidocephalosporanic acid in the form of the potassium salt, having a maximum in its ultraviolet absorption spectrum at 257 m (6:7850).

EXAMPLE 7-(N'-pyrrylacetamid0) cephalospora-nic acid 7-aminocephalosporanic acid (2.2 g.), N-pyrroleacetic acid (1.0 g., prepared by the method of Clemo et al., J. Chem. Soc., 1931, 49), and dicyclohexylcarbodiimide (1.7 g.) were reacted in tetrahydrofuran generally according to the procedure of Example 14. The residue obtained by evaporation of the ethyl acetate extract was dissolved in 20 ml. of ethanol, and to the solution was added 0.5 g. of potassium acetate in 10 ml. of ethanol. The resulting precipitate was collected and dried. The yield was 300 mg. of 7-(N'-pyrrylacetamido)cephalosporanic acid in the form of the potassium salt, having a maximum in its ultraviolet absorption spectrum at 258 mu (e:7670).

I claim:

7-5-(2'-furyl)propionamidocephalosporanic acid. 7-(2'-fury1)acetamidocephalosporanic acid. 7-(.2'-thienyl) acetamidocephalosporanic acid. 7-('y-2'-thienyl-n-butyramido)cephal0sporan ic acid. 7-(1-pyrazoleacetamido)cephalosporanic acid. 7-(N'-pyrrylacetamido)cephalosporanic acid. The antibiotic substances of the class represented by the following formula wherein R is C -C alkylene; n is a member of the class consisting of 0 and l; and R is a heteromonocyclic radical selected from the group consisting of (A) dioxanyl, furyl, imidazolyl, isoxazolyl, morpholinyl, oxazolyl, pyranyl, pyrazinyl, pyrazolyl, pyridyl, pyrimidyl, pyrryl, thiazolyl, thienyl, triazinyl, and triazolyl; (B) tetrahydrofuryl, imidazolinyl, imidazolidyl, piperidyl, tetrahydropyrimidyl, and pyrrolidyl; and (C) the members of subgroups A and B having attached to the ring thereof at least one member of the class consisting of chlorine, fluorine, bromine, iodine, nitro, methoxy, and C -C alkyl; and the salts thereof with pharmaceutically acceptable cations.

8. 7-(2'-thienyl)acetamidocephalosporanic acid sodium salt.

References Cited by the Examiner Bergman: The Chemistry of Acetylene and Related Compounds, page (1948).

Burger: Medicinal Chemistry, pages 46-47 (1960).

Morton: The Chemistry of Heterocyclic Compounds, page VI of the preface (1946).

NICHOLAS S. RIZZO, Primary Examiner. 

7. THE ANTIBIOTIC SUBSTANCES OF THE CLASS REPRESENTED BY THE FOLLOWING FORMULA 